The recent finding that presynaptic stimulation can result in the corelease of different neurotransmitters from the same neuron provides an important and novel intrasynaptic mechanism by which synaptic transmission may be modulated. The objective of the proposed research is to study the biochemical mechanisms which regulate corelease and which determine neuronal output following presynaptic stimulation. The experiments will be performed on the cholinergic nerve terminals of the richly innervated Torpedo electric organ which, as we have recently shown, contain an opioid like peptide (OLP) which is coreleased with acetylcholine (ACh). The homogeneity of this preparation, and its endowment with presynaptic muscarinic and opiate receptors whose activation regulate release, render it uniquely suited for the proposed study. The Torpedo OLP will be purified and characterized. Its corelease with ACh will be studied by determining the dependency of the release of ACh and OLP from isolated nerve terminals ('synaptosomes') and from tissue slices on the mode and extent of stimulation, and by comparing their metabolism within the nerve terminal (e.g. synthesis, uptake, storage, compartmentation). The regulation of corelease by presynaptic muscarinic and opiate receptors will be studied by determining their relative effects on the release of ACh and OLP and by investigating the biochemical mechanisms underlying these effects. This will be pursued by extending our preliminary findings that activation of the opiate receptor inhibits release by blocking the influx of Ca2+ into the nerve terminal and that the muscarinic effects on release are mediated by a PGE2-like prostaglandin which is its second messenger and inhibits release by interference with the coupling between intraterminal Ca2+ and release. The proposed study of corelease and of the regulation of synaptic transmission by interacting neurotransmitters is expected to further our understanding of the function of the normal synapse and to yield conceptual and experimental tools which will be applicable to the study of synaptic dysfunction.